不同产地梅花鹿鹿茸药材中5种核苷类成分的含量测定及聚类分析

　　摘 要 目的：建立同時测定梅花鹿鹿茸药材中尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷 5 种核苷类成分含量的方法，并比较不同产地梅花鹿鹿茸药材中上述核苷类成分含量的差异。方法：采用高效液相色谱法。色谱柱为Agilent Eclipse Plus C18，流动相为甲醇-0.07%冰醋酸溶液（4 ∶ 96，V/V），流速为1.0 mL/min， 检测波长为254 nm，柱温为28 ℃，进样量为5 μL。采用SPSS 19.0软件对30批不同产地梅花鹿鹿茸药材（S1～S30）进行聚类分析。结果：尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷的检测质量浓度线性范围均为0.001～0.01 mg/mL（r均为0.999 9）；定量限分别为1.489 1、1.927 9、4.880 9、7.884 6、8.092 1 ng，检测限分别为0.446 7、0.578 4、1.464 3、2.365 4、2.427 7 ng；精密度、稳定性、重复性试验的RSD均小于2%；加样回收率分别为99.69%～103.65%（RSD=1.40%，n=9）、97.77%～103.26%（RSD=1.67%，n=9）、97.82%～101.81%（RSD=1.12%，n=9）、99.30%～104.82%（RSD=1.72%，n=9）、98.13%～100.20%（RSD=0.64%，n=9）。30批药材样品中尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷的含量存在差异。上述药材样品可聚为3大类，即S16～S18、S22～S30聚为一类，S8～S15、S19～S21聚为一类，S1～S7聚为一类。结论：该方法精密度、稳定性、重复性均较好，可用于同时测定梅花鹿鹿茸药材中尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷的含量；梅花鹿鹿茸药材的质量受生态环境、养殖技术等因素的影响，存在较大差异。
　　关键词 梅花鹿鹿茸；尿嘧啶；次黄嘌呤；尿苷；肌苷；鸟苷；高效液相色谱法；含量测定；聚类分析
　　中图分类号 R284.1 文献标志码 A 文章编号 1001-0408（2018）14-1945-05
　　DOI 10.6039/j.issn.1001-0408.2018.14.17
　　ABSTRACT OBJECTIVE： To set up a method for simultaneous determination of 5 nucleosides as uracil， hypoxanthine， uridine， creatinine and guanosine in Cervus nippon， to compare the differences of above nucleosides content in C. nippon among different producing areas. METHODS： HPLC method was adopted. The determination was performed on Aglient Eclipse Plus C18 column with methanol-0.07% glacial acetic acid （4 ∶ 96，V/V） as mobile phase at the flow rate of 1.0 mL/min. The detection wavelength was 254 nm. The column temperature was set at 28 ℃， and sample size was 5 ?L. Cluster analysis was conducted for 30 batches of C. nippon from different producing areas by using SPSS 19.0 software. RESULTS： The linear ranges of uracil， hypoxanthine， uridine， creatinine and guanosine were 0.001-0.01 mg/mL （all r≥0.999 9）. The limits of quantitation were 1.489 1， 1.927 9， 4.880 9， 7.884 6， 8.092 1 ng， and the limits of detection were 0.446 7， 0.578 4， 1.464 3， 2.365 4， 2.427 7 ng， respectively. RSDs of precision， stability， reproducibility tests were lower than 2%. The recoveries were 99.69%-103.65%（RSD=1.40%，n=9），97.77%-103.25%（RSD=1.67%，n=9）， 97.82%-101.81%（RSD=1.12%， n=9）， 99.30%-104.82%（RSD=1.72%，n=9） and 98.13%-100.20%（RSD=0.64%，n=9）. The contents of uracil， hypoxanthine， uridine， creatinine and guanosine were significantly different in 30 batches of C. nippon， which were clustered into 3 categories. S16-S18 and S22-S30 were clustered into one category； S8-S15 and S19-S21 were clustered into one category； S1-S7 were clustered into one category. CONCLUSIONS： The method has good precision， stability and reproducibility. It is suitable for simultaneous determination of uracil， hypoxanthine， uridine， creatinine and guanosine in C. nippon. The quality of C. nippon has great difference due to ecological environment and technical factors of farming.

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